Transgenic plant having herbicide resistance
a technology of herbicide resistance and transgenic plants, which is applied in the field of transgenic plants, can solve the problems of difficult identification of the molecular species of gst that metabolizes a particular drug, and the registration of pyroxasulfone, and achieve the effect of stably cultivating and producing
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example 1
[0158]In Example 1, the GST identified under Accession Code: Q8GTC0 (referred to as “TaQ8GTC0” herein) among GSTs in wheat plants was analyzed in terms of the activity of glutathione conjugation to VLCFAE-inhibiting herbicides.
E. coli>
[0159]FIG. 1 shows a constitution of a construct of the TaQ8GTC0 protein expressed in E. coli. At the outset, cDNA was synthesized via reverse transcription from RNA prepared from the shoots of wheat (Nohrin No. 61) as a template. The cDNA obtained was used as a template to perform PCR with the use of a set of primers (TaQ8GTC0-H (SEQ ID NO: 3) and TaQ8GTC0-D (SEQ ID NO: 4)). Thus, a fragment of the TaQ8GTC0 gene comprising the NdeI recognition site at the 5′ terminus and the BamHI recognition site at the 3′ terminus was obtained.
[0160]Subsequently, the TaQ8GTC0 gene fragment was digested with NdeI and BamHI, the digested TaQ8GTC0 gene fragment was designated as an insert, and pET22b (+) that was also treated with NdeI and BamHI was designated as a ...
example 2
[0169]In Example 2, transgenic rice plants comprising the TaQ8GTC0 gene introduced therein were prepared, and sensitivity of the transgenic rice plants to VLCFAE-inhibiting herbicides was tested.
[0170]In Example 2, a vector comprising the TaQ8GTC0 gene for rice transformation (R-5-TaQ8GTC0) was prepared, as shown in FIG. 5.
[0171]Specifically, cDNA was first synthesized via reverse transcription from RNA prepared from the shoots of wheat (Nohrin No. 61) as a template. The cDNA obtained was used as a template to perform PCR with the use of a set of primers (TaQ8GTC0-M (SEQ ID NO: 5) and TaQ8GTC0-N(SEQ ID NO: 6)). Thus, a fragment of the TaQ8GTC0 gene comprising the SalI recognition site at the 5′ terminus and the NotI recognition site at the 3′ terminus was obtained.
[0172]Subsequently, the TaQ8GTC0 gene fragment was digested with SalI and NotI, the treated TaQ8GTC0 gene was designated as an insert, and pENTR-1A (Thermo Fisher Scientific Inc.) that was also digested with SalI and NotI ...
example 3
[0189]In Example 3, transgenic Arabidopsis thaliana plants comprising the TaQ8GTC0 gene introduced therein were produced and sensitivity of the transgenic Arabidopsis thaliana plants to VLCFAE-inhibiting herbicides was tested.
Arabidopsis thaliana Plants>
[0190]At the outset, as shown in FIG. 8, a vector for Arabidopsis thaliana transformation comprising the TaQ8GTC0 gene (A-3-TaQ8GTC0) was produced. Specifically, the entry clone comprising the TaQ8GTC0 gene (ORF) inserted into a site between attL1 and attL2 of pENTR-1A (Thermo Fisher Scientific Inc.) (pENTR1A-TaQ8GTC0, KLB-649) was used in the LR reaction with the destination vector (PalSelect A-3, Inplanta Innovations Inc.) to produce a vector for Arabidopsis thaliana transformation. This vector for Arabidopsis thaliana transformation was produced by inserting the TaQ8GTC0 gene into the attB sequence of PalSelect A-3. Subsequently, the vector for Arabidopsis thaliana transformation was introduced into the E. coli HST02 strain via tr...
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